Lens Disulfide crosslink - Evidence of highly conserved β-crystallin disulfidome that can be mimicked by in vitro oxidation in age-related human cataract and glutathione depleted LEGSKO mouse lens

ABSTRACT: Low glutathione levels are associated with crystallin oxidation in age-related nuclear cataract (ARNC). To understand the role of cysteine residue oxidation, we used the novel approach of comparing human cataracts with glutathione-depleted LEGSKO mouse lenses for intra- vs. intermolecular disulfide crosslinks using 2D-PAGE and proteomics, and then systematically identified in vivo and in vitro all disulfide forming sites using ICAT labeling method coupled with proteomics. Crystallins rich in intramolecular disulfides were abundant at young age in human and WT mouse lens but shifted to multimeric intermolecular disulfides at older age. The shift was ~4x accelerated in LEGSKO lens. Most cysteine disulfides in β-crystallins (except βA4 in human) were highly conserved in mouse and human and could be generated by oxidation with H2O2, while γ-crystallin oxidation selectively affected γC23/42/79/80/154, γD42/33 and γS83/115/130 in human cataracts, and γB79/80/110, γD19/109, γF19/79, γE19, γS83/130 and γN26/128 in mouse. Analysis based on available crystal structure suggests that conformational changes are needed to expose C42, C79/80, C154 in γC; C42, C33 in γD, and C83, C115 and C130 in γS. In conclusion, while the β-crystallin disulfidome is highly conserved in ARNC and LEGSKO mouse, and reproducible by in vitro oxidation, some of the disulfide formation sites in γ-crystallins necessitate prior conformational changes. Overall, the LEGSKO mouse model is closely reminiscent of ARNC.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Lens Nucleus

DISEASE(S): Nuclear Senile Cataract

SUBMITTER: Xingjun Fan  

LAB HEAD: Xingjun Fan

PROVIDER: PXD002658 | Pride | 2015-10-16

REPOSITORIES: Pride